To determine the molecular basis for the distinct biological responses of platelet-derived growth factor A chain (PDGF-A) in NIH/3T3 cells, a chimerae consisting of the extracellular domain of the human colony stimulating factor receptor (CSF-1R) (fms) linked to the cytoplasmic domain of the aPDGFR (aR) containing a series of deletion or point mutations was constructed. The ability of fms/aR chimerae to mediate CSF-1 dependent anchorage-independent growth, focus formation and chemotaxis of NIH/3T3 cells was then examined. The results provide evidence that a domain encompassing amino acid residues 977-1024 of the aPDGFR is required for ligand-dependent focus formation but not chemotaxis or anchorage-independent growth, and that tyrosine residues within this domain constitute the major binding site for phospholipase g (PLCg). Thus, these findings suggest that the focus-forming function of aPDGFR correlates well with the ability of the receptor to bind PLCg. In contrast to the mutations within the carboxy-terminal domain, a double mutation or a deletion within the kinase insert domain completely abrogated receptor-associated PI-3 kinase and fully inhibited ligand-stimulated cell movement but not focus formation or colony formation of NIH/3T3 transfectants. These findings indicate that the ligand-dependent association of the aPDGFR with PI-3 kinase activity is required for chemotaxis, but not growth or focus formation. In summary, this data has defined the molecular basis for PDGF-induced focus formation and chemotaxis indicating that the mechanism of focus formation may be distinguished from those of chemotaxis or anchorage-independent growth.